Characterization of the pTFI91 -family replicon of Thiobacillus ferrooxidans plasmids

Abstract

Plasmids found in six strains of Thiobacillus ferrooxidans were mapped and compared in an effort to detect the origin of replication. Four strains yielded an identical 9.8-kb plasmid, pTFI91. Restriction mapping and Southern blot hybridization analysis were used to confirm this finding. Dissimilar plasmids found in two other strains contained a conserved 2.2-kb SacI region common to pTFI91. DNA sequence analysis of this region showed structural features common to bacterial plasmid replicons. A comparison of the pTFI91 origin with those of T. ferrooxidans pTF-FC2 and other broad host range vectors did not show significant homologous DNA sequences. To verify the replication function, a chloramphenicol acetyl transferase marker gene was ligated at the unique sites of pTFI91, and the plasmid was transformed into Escherichia coli DH5 alpha cells but no transformants were identified. To test the replication of pTFI91 independent of DNA polymerase I in E. coli, different restriction fragments of pTFI91 were cloned into pHSG398 (Cmr, ColEI origin) and transformed into the polA1 mutant SF800, but chloramphenicol-resistant transformants were not detected. Electrotransformation of T. ferrooxidans TFI-70 and Pseudomonas putida ATCC 19151 also failed to yield transformants. The results suggested that the pTFI91 plasmid replicon does not function either in E. coli or in P. putida. Since pTFI91 contains the same origin of replication as other plasmids in several other T. ferrooxidans strains, this replicon may be commonly distributed in T. ferrooxidans.

Metal resistance in Acidocella strains and plasmid-mediated transfer of this characteristic to Acidiphilium multivorum and Escherichia coli

“…Probably, plasmids of these strains contained stretches of homologous regions or similar structural motifs related to some of their common physiological functions. It should be mentioned that T. ferrooxidans strains isolated from different sites contain plasmids which have highly homologous regions (3,28).…”

“…Genetic inheritance of metal resistance is often located on plasmids (29,32), but for acidophiles such a list awaits generation. Because of the importance of plasmid-borne metal resistance in genetic manipulation of leaching microbes (24,25) and due to the very low to nonexistent level of expression of this characteristic in heterologous systems (18), endogenous plasmids carrying genes for this phenotypic trait in T. ferrooxidans or other acidophiles may find wide application in the development of bioleaching operations (3,28). Since many acidophilic bacterial strains contain plasmids (8,15,16,21), there is a high probability that a few of them harbor metal resistance-conferring plasmids.…”

Two Large, Related, Cryptic Plasmids from Geographically Distinct Isolates of Sulfobacillus thermotolerans

“…The finding that backbones of plasmids isolated from different strains of S. thermotolerans from different geographical locations are related but that the accessory genes are different appears to be a recurring theme in iron-and sulfuroxidizing bacteria. Species-specific plasmid backbones have been reported in Acidithiobacillus ferrooxidans (7,12,43), and Acidithiobacillus caldus (44). In A. caldus and S. thermotolerans, these plasmids appear to be either mobilizable or conjugable and may be involved in the movement of the accessory genes within isolates of the same species and possibly also between species.…”

A geographically widespread plasmid from Thiobacillus ferrooxidans has genes for ferredoxin-, FNR-, prismane- and NADH-oxidoreductase-like proteins which are also located on the chromosome

“…Several authors have noted that some plasmids found in T. ferrooxidans have a very wide geographic distribution (Sanchez et al , 1986; Valenti et al , 1989, 1990; Chakravarty et al , 1995). Members of one widely distributed T. ferrooxidans plasmid family share a highly conserved replicon, part of which is present on a 2.2 kbp Sac I fragment (Chakravarty et al , 1995).…”

“…Several authors have noted that some plasmids found in T. ferrooxidans have a very wide geographic distribution (Sanchez et al , 1986; Valenti et al , 1989, 1990; Chakravarty et al , 1995). Members of one widely distributed T. ferrooxidans plasmid family share a highly conserved replicon, part of which is present on a 2.2 kbp Sac I fragment (Chakravarty et al , 1995). One particular T. ferrooxidans plasmid has been isolated by workers in three different laboratories and although the estimates of its size vary slightly (19.6–20.0 kbp), the plasmids appear to have an identical restriction enzyme map to pTF35 (Rawlings et al , 1983).…”